Binding of the RamR Repressor to Wild-Type and Mutated Promoters of the
            <i>ramA</i>
            Gene Involved in Efflux-Mediated Multidrug Resistance in Salmonella enterica Serovar Typhimurium

Baucheron, Sylvie; Coste, Franck; Canepa, Sylvie; Maurel, Marie‐Christine; Giraud, Étienne; Culard, Françoise; Castaing, B.; Roussel, Alain; Cloeckaert, Axel

doi:10.1128/aac.05444-11

Cited by 45 publications

(40 citation statements)

References 24 publications

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“…As shown in Fig. 4a, the shift of the DNA band was dependent on the protein concentration 16 . Addition of berberine, crystal violet, dequalinium, ethidium bromide or rhodamine 6G to the RamR-DNA complex resulted in the loss of the retarded band, indicating the separation of the protein and DNA components.…”

Section: Structure Of the Ramr Proteinmentioning

confidence: 92%

“…Bacterial multidrug efflux regulators usually control the expression of multidrug efflux pumps by binding to inverted repeat sequences in their promoter regions. A recent study using footprinting analysis revealed the 28-bp RamR-binding site 16 . On the basis of the RamR footprint and electrophoretic mobility shift assays (EMSAs), it was proposed that RamR interacts with P ramA as a dimer of dimers 16 .…”

Section: Structure Of the Ramr Proteinmentioning

confidence: 99%

“…A recent study using footprinting analysis revealed the 28-bp RamR-binding site 16 . On the basis of the RamR footprint and electrophoretic mobility shift assays (EMSAs), it was proposed that RamR interacts with P ramA as a dimer of dimers 16 . Our results suggest that the binding of multiple drugs to RamR may reduce its DNA-binding affinity, resulting in the induction of ramA expression.…”

Section: Structure Of the Ramr Proteinmentioning

confidence: 99%

“…RamR binds to the region upstream of ramA. Its binding site covers essential features of the ramA promoter, including the À 10 conserved region, the transcriptional start site of ramA and two 7-bp inverted repeats 16 . Various types of mutations in ramR and the ramR-ramA intergenic region were identified in multidrug-resistant strains of S. typhimurium, other S. enterica serovars and K. pneumoniae, which result in increased expression of ramA and an increase in efflux-mediated multidrug resistance 15,17,18 .…”

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confidence: 99%

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The crystal structure of multidrug-resistance regulator RamR with multiple drugs

et al. 2013

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RamR is a transcriptional repressor of the gene-encoding RamA protein, which controls the expression of the multidrug efflux system genes acrAB-tolC. RamR is an important multidrug-resistance factor, however, its structure and the identity of the molecules to which it responds have been unknown. Here, we report the crystal structure of RamR in complex with multiple drugs, including berberine, crystal violet, dequalinium, ethidium bromide and rhodamine 6G. All compounds are found to interact with Phe155 of RamR, and each compound is surrounded by different amino acid residues. Binding of these compounds to RamR reduces its DNA-binding affinity, which results in the increased expression of ramA. Our results reveal significant flexibility in the substrate-recognition region of RamR, which regulates the bacterial efflux participating in multidrug resistance.

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Section: Structure Of the Ramr Proteinmentioning

confidence: 92%

Section: Structure Of the Ramr Proteinmentioning

confidence: 99%

Section: Structure Of the Ramr Proteinmentioning

confidence: 99%

mentioning

confidence: 99%

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The crystal structure of multidrug-resistance regulator RamR with multiple drugs

et al. 2013

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“…Binding at a similar promoter-proximal site has been shown recently for another TetR-family regulator, RamR, of Salmonella enterica (4), in contrast to other family members that have been shown to bind at a location more distal to the transcriptional start (33,35). However, these repressors have all been shown to bind at a single site to mediate repression of the operon as either a dimer or a dimer of dimers (4,33,35), whereas CifR binds at two distinct operators, separated by 28 bp, to control the expression of divergently transcribed loci. The auto-regulation exhibited by CifR, common in the TetR family, is believed to be a feedback control mechanism that ensures optimal repressor concentration (33).…”

Section: Discussionmentioning

confidence: 99%

Epoxide-Mediated CifR Repression of cif Gene Expression Utilizes Two Binding Sites in Pseudomonas aeruginosa

et al. 2012

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bPseudomonas aeruginosa secretes an epoxide hydrolase virulence factor that reduces the apical membrane expression of ABC transporters such as the cystic fibrosis transmembrane conductance regulator (CFTR). This virulence factor, named CFTR inhibitory factor (Cif), is regulated by a TetR-family, epoxide-responsive repressor known as CifR via direct binding and repression. We identified two sites of CifR binding in the intergenic space between cifR and morB, the first gene in the operon containing the cif gene. We have mapped these binding sites and found they are 27 bp in length, and they overlap the ؊10 and ؉1 sites of both the cifR and morB regulatory region and the start of transcription, respectively. In addition, we found that CifR binds to each repression site with differing affinity. Mutagenesis of these binding sites resulted in a loss of DNA binding in vitro, and mutation of one of these sites in vivo resulted in an increase in transcription of both the cif and cifR genes. We characterized cif and cifR gene expression in sputum and found that, whereas cif gene expression varied relative to an in vitro coculture control, cifR gene expression was consistently higher. Analysis of a longitudinal sample of CF isolates from nine patients revealed that Cif protein was expressed over time, although variably, and these changes could not be linked to mutations in the cifR gene or the promoters of these genes. Finally, we tested CifR responsiveness to other epoxides and showed that CifR can respond to multiple epoxides to various degrees.

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TetR and OmpR family regulators in natural product biosynthesis and resistance

Patil,

Sharma,

Bhaskarwar

et al. 2023

Proteins

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This article provides a comprehensive review and sequence‐structure analysis of transcription regulator (TR) families, TetR and OmpR/PhoB, involved in specialized secondary metabolite (SSM) biosynthesis and resistance. Transcription regulation is a fundamental process, playing a crucial role in orchestrating gene expression to confer a survival advantage in response to frequent environmental stress conditions. This process, coupled with signal sensing, enables bacteria to respond to a diverse range of intra and extracellular signals. Thus, major bacterial signaling systems use a receptor domain to sense chemical stimuli along with an output domain responsible for transcription regulation through DNA‐binding. Sensory and output domains on a single polypeptide chain (one component system, OCS) allow response to stimuli by allostery, that is, DNA‐binding affinity modulation upon signal presence/absence. On the other hand, two component systems (TCSs) allow cross‐talk between the sensory and output domains as they are disjoint and transmit information by phosphorelay to mount a response. In both cases, however, TRs play a central role. Biosynthesis of SSMs, which includes antibiotics, is heavily regulated by TRs as it diverts the cell's resources towards the production of these expendable compounds, which also have clinical applications. These TRs have evolved to relay information across specific signals and target genes, thus providing a rich source of unique mechanisms to explore towards addressing the rapid escalation in antimicrobial resistance (AMR). Here we focus on the TetR and OmpR family TRs, which belong to OCS and TCS, respectively. These TR families are well‐known examples of regulators in secondary metabolism and are ubiquitous across different bacteria, as they also participate in a myriad of cellular processes apart from SSM biosynthesis and resistance. As a result, these families exhibit higher sequence divergence, which is also evident from our bioinformatic analysis of 158 389 and 77 437 sequences from TetR and OmpR family TRs, respectively. The analysis of both sequence and structure allowed us to identify novel motifs in addition to the known motifs responsible for TR function and its structural integrity. Understanding the diverse mechanisms employed by these TRs is essential for unraveling the biosynthesis of SSMs. This can also help exploit their regulatory role in biosynthesis for significant pharmaceutical, agricultural, and industrial applications.

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Binding of the RamR Repressor to Wild-Type and Mutated Promoters of the ramA Gene Involved in Efflux-Mediated Multidrug Resistance in Salmonella enterica Serovar Typhimurium

Cited by 45 publications

References 24 publications

The crystal structure of multidrug-resistance regulator RamR with multiple drugs

The crystal structure of multidrug-resistance regulator RamR with multiple drugs

Epoxide-Mediated CifR Repression of cif Gene Expression Utilizes Two Binding Sites in Pseudomonas aeruginosa

TetR and OmpR family regulators in natural product biosynthesis and resistance

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